One of the major factors contributing to the high fatality of pancreatic cancer is the poor response of most pancreatic cancer patients to therapies including radiation therapy and chemotherapy. Our long- term objective of this project is to understand the molecular mechanisms underlying the resistance of pancreatic cancer to therapies. Induction of apoptosis in target cells is a key mechanism by which radiation and chemotherapy induces cell killing. Mitochondrion and mitochondrion-generated reactive oxygen species (ROS) play essential roles in the cell death pathways. In the mitochondria dependent apoptotic pathways, after the release of the apoptotic factors, such as cytochrome c, the downstream biochemical pathways appear to be shared by different cells. We hypothesize that the mitochondrial proteins involved in ROS production or scavenging, which determine the ROS levels in mitochondria, may be responsible for the resistance of pancreatic cells to therapies. Accordingly, we propose to use quantitative proteomic methods to systematically screen mitochondrial proteins that are involved in ROS production and scavenging in pancreatic cancer cells with different sensitivity to ROS inducing drugs (Specific Aim 1). After the proteomic screening, RNAi or overexpression techniques will be used to manipulate the expression of the identified target proteins in cells, and the effects of the altered expression of the target proteins on cellular ROS accumulation, and cellular apoptosis will be determined (Specific Aim 2). Identifications of the protein factors that are closely associated with the resistance of pancreatic cancer cells to anti-cancer drugs may provide valuable information for designing new novel therapeutic strategies to overcome the resistance of pancreatic cancer to therapies. PUBLIC HEALTH RELEVANCE: The objective of this project is to identify novel protein targets that may be responsible for chemo- resistance of pancreatic cancer. Novel quantitative proteomic methods will be used to systemically compare mitochondrial redox proteins in chemo-sensitive and chemo-resistant pancreatic cancer cells. Molecular and cell biological techniques will be used to assess the biological functions of the identified target proteins.